The embryonic epsilon-globin gene is the first member of the beta-globin family of genes to be expressed during development. Expression of the beta-like-globin genes is under the control of the beta-globin locus control region (LCR), encompassing erythroid specific DNase I hypersensitive sites (HS) 5-18 kilobases upstream of the epsilon-globin gene. The LCR has long range effects on the chromatin structure of the beta-globin locus, and it activates transcription of the different globin gene promoters as the genes are expressed sequentially during development. Much of our current understanding of the process of gene activation has been gained from in vitro studies where DNA is not assembled with histones and other chromosomal proteins as it is in the nucleus. To study the activation of a gene in its natural context of chromatin in vivo we have used minichromosomes containing the human epsilon-globin gene and portions of the beta globin LCR enhancer. Expression of hue minichromosomal epsilon-globin gene in K562 human erythroid cells is completely dependent on the presense if the beta- globin LCR in cis. Using restriction endonuclease access, and indirect end labelling experiments, we have observed that the promoter and 5' proximal structural regions of the minichromosomal epsilon-globin gene are covered by an array of positioned nucleosomes. Both types of studies suggest alteration or loss of a single promoter-proximal nucleosome upon activation of the gene for transcription. Regulated expression of the epsilon-globin gene on the minichromsomes offers a means too study at high resolution, the promoter-enhancer interaction of an activated gene assembled in vivo into chromatin.